Journal of Chromatography A, 1395 (2015) 41–47
Contents lists available at ScienceDirect
Journal of Chromatography A journal homepage: www.elsevier.com/locate/chroma
Determination of volatile organic compounds in water using headspace knotted hollow fiber microextraction Pai-Shan Chen a,b,∗ , Yu-Hsiang Tseng a , Yuh-Lin Chuang a , Jung-Hsuan Chen a,b a b
Department and Graduate Institute of Forensic Medicine, National Taiwan University, Taipei 10002, Taiwan Forensic and Clinical Toxicology Center, National Taiwan University College of Medicine and National Taiwan University Hospital, Taiwan
a r t i c l e
i n f o
Article history: Received 10 January 2015 Received in revised form 24 March 2015 Accepted 24 March 2015 Available online 31 March 2015 Keywords: Headspace Hollow fiber microextraction Volatile organic compounds Gas chromatography–mass spectrometry River water Wastewater
a b s t r a c t An efficient and effective headspace microextraction technique named static headspace knotted hollow fiber microextraction (HS-K-HFME) has been developed for the determination of volatile organic compounds (VOCs) in water samples. The knot-shaped hollow fiber is filled with 25 L of the extraction solvent. The excess solvent forms a large droplet (13 L) and is held in the center of the knot. Even after 20 min of extraction time at high temperature (95 ◦ C) without cooling, there was still enough volume of extraction solvent for gas chromatography–mass spectrometry (GC–MS) analysis, which extends the choice of solvents for headspace LPME. Moreover, the knot-shaped fiber has a larger extraction contact interface, which increases the rate of mass transfer between the headspace and extraction solvent film attached to the fiber, thus improving the extraction efficiency. The effects of extraction solvent, temperature, stirring rate, salt concentration and extraction time on extraction performance were optimized. The calibration curves exhibited coefficients of determination (R2 ) ranging from 0.9957 to 0.9999 and the limit of detection (LOD) ranged from 0.2 to 10 g L−1 . Relative standard deviations (RSDs) ranged from 4.5% to 11.6% for intraday measurements (n = 5). Interday (n = 15) values were between 2.2% and 12.9%. The relative recoveries (RRs) ranged from 90.3% to 106.0% for river water and 95.9% to 103.6% for wastewater. © 2015 Elsevier B.V. All rights reserved.
1. Introduction Volatile organic compounds (VOCs) are defined as having a boiling point that ranges between 50 ◦ C and 260 ◦ C [1]. VOCs are widely emitted from sources such as paints, varnishes, solvents and preservatives, and become major environmental pollutants because of their high volatility and toxicity [2,3]. Due to their physical properties, VOCs can easily cross lipid membranes and distribute to well-perfused organs; exposure to VOCs may thus result in both acute and chronic health effects. Diethyl ether (DE) and ethyl acetate (EA) are two common VOCs, which are used worldwide in flavoring and perfumery and in smokeless power manufacture. They are mildly irritating to eyes, nose and throat. Exposure to high levels may cause dizziness or to pass out. Especially for DE, severe over exposure may result in death [4]. Certain VOCs are more toxic and persistent in water, soil, and organisms, such as dichloromethane (DCM), toluene, o-xylene, m-xylene, and
∗ Corresponding author at: Department and Graduate Institute of Forensic Medicine, National Taiwan University, Taipei 10002, Taiwan. Tel.: +886 2 2312 3456x65495; fax: +886 2 2321 8438. E-mail address: [email protected] (P.-S. Chen). http://dx.doi.org/10.1016/j.chroma.2015.03.067 0021-9673/© 2015 Elsevier B.V. All rights reserved.
p-xylene. They are considered one of the major causes of environmental pollution because of widespread occurrences of leakage from underground gasoline storage tanks and spills. Long term exposure of them has been associated with liver and kidney damage, intestinal tract disturbances and central nervous system depression [4–6]. Therefore, the United States Environmental Protection Agency (EPA) has regulated 0.005 and 10 mg L−1 as the maximum permissible contaminant level for dichloromethane and total xylene respectively in drinking water [7]. As a result of this low limit, it is necessary to develop highly sensitive and efficient analytical methods to detect VOCs in the aquatic environment. Analyte extraction and pretreatment is the most challenging and time-consuming step in most chromatographic procedures. In the last few years, research trends in separation science have oriented toward minimizing the sample pretreatment steps. Solidphase microextraction (SPME) and liquid-phase microextraction (LPME) have been developed based on this concept. SPME was first introduced by Pawliszyn and coworkers [8] and involves the partition of analytes between sample matrices and a polymercoated stationary phase on a silica fiber. It enables the extraction and simultaneous preconcentration of analytes from aqueous samples. There are three major types of SPME: direct-immersion [9], headspace (HS) [10–12] and membrane-protected SPME [13].
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P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47
Fig. 1. The extraction equipment of HS-K-HFME.
Among them, HS-SPME analysis is the most common technique for the determination of VOCs as it significantly reduces the interference from dirty matrices, which is generally associated with direct-immersion SPME [14–16]. Lately, LPME has emerged as an attractive alternative to conventional liquid–liquid extraction (LLE). Jeannot and Cantwell proposed single-drop microextraction (SDME) in which an 8-L droplet of organic solvent was suspended at the end of a Teflon rod and immersed in an aqueous solution to extract analyte specimens [17]. An improvement involved the use of a microsyringe to hold the solvent drop [18]. To increase the rate of mass transfer, Lee et al. developed a dynamic liquid phase microextraction technique using a short hollow fiber inserted onto a microsyringe [19,20]. The hydrophobic and porous fiber was filled with organic solvent, which stabilizes the solvent successfully in comparison with holding a droplet at the tip of the microsyringe [21–23]. The immersion mode yields high enrichment factors, high sensitivity and ruggedness. However, headspace LPME using a hollow fiber presents more challenges than other types of LPME because the extraction solvents compatible with gas chromatography (GC) usually evaporate quickly in the headspace. For headspace LPME, SDME and fiber-protected LPME have been developed. Theis et al. [24] introduced headspace SDME using 1octanol as the extraction solvent for analyzing VOCs, which has a relatively low vapor pressure. Shen and Lee [25] used dynamic headspace LPME to improve the stability of headspace SDME. A thin organic solvent film was formed on the inner syringe wall. The dynamic operation increased and refreshed the interface between the gas phase and the extraction solvent, thus increasing the extraction efficiency significantly. Jiang et al. [26] developed a dynamic hollow fiber-supported headspace liquid-phase microextraction (DHF-HS-LPME) technique. A hollow fiber filled with the solvent was inserted onto a microsyringe needle which was kept vertically in the headspace of the sample solution. The solvent within the fiber could be moved using a programmable syringe pump. This method enabled the use of higher solvent volumes, increased the extraction contact interface and stabilized the organic solvent within the fiber. In such analyses, headspace gas is usually injected into a GC directly. A major concern is that the organic solvents commonly used in GC have high vapor pressures, which result in quick evaporation in the headspace and limits the possible selection
of extraction solvents. In 2007, Huang and Huang [21] applied a solvent cooling system to the microsyringe which decreased the temperature of the solvent when it was drawn into the syringe. This decreased temperature minimized the loss of extraction solvent but also caused slow equilibration and led to longer extraction times. Chen et al. [27] developed a dynamic hook-type liquid-phase microextraction (DHT-LPME). The extraction solvent was easily and completely withdrawn into the microsyringe, while the hook shape removed interference from air bubbles which commonly persist when the hollow fiber is placed vertically. Subsequently the same group introduced dynamic headspace time-extended helix LPME (DHS-TEH-LPME) to overcome the slow equilibrium caused by cooling system. An aluminum heating block was used to heat the sample vial while the extraction solvent was condensed using a cooling system. The extraction temperature up to 80 ◦ C was used for 60 min. Chen et al. [28] developed a PTFE vial cap with a cylindrical cavity to hold a 40-L droplet of volatile extraction solvents, such as acetone, in the headspace. The cooling system, based on a thermoelectric cooler (TEC), was used to lower the temperature of the vial cap below zero. This method enabled a limit of detection (LOD) for chlorobenzenes between 4 and 8 g L−1 . It is worth noting that, in order to extend the selection of solvents and reduce solvent loss during headspace, LPME extraction processes, a cooling system is usually required. However, this also results in slow equilibrium and longer extraction times to obtain satisfactory extraction efficiency. To address these issues, we proposed a simple and efficient headspace LPME technique named headspace knotted hollow fiber microextraction (HS-K-HFME) for the determination of VOCs in water samples. A 2.5 cm long hollow fiber was bent into a figure eight knot shape (Fig. 1) and filled with 25 L of extraction solvent. AA 2.5 cm fiber can hold 12 L of solvent approximately. The excess solvent which was not held inside the fiber formed a large droplet (approximately 13 L). The droplet was held steadily at the center of the knot. As a result of the figure eight shape, there is no interference from air bubbles in the fiber. The extraction solvent is withdrawn into the microsyringe easily and the long fiber has a large contact surface that increases the extraction interface between the headspace and the organic solvent. This large volume is sufficient to compensate for evaporative losses during sampling. Moreover, during the extraction of VOCs, the specimen solution can also be heated.
P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47 Table 1 Qualitative GC–MS data for the studied VOCs. Analyte
Quantitative and confirmed ions (m/z)
Diethyl ether (DE) Ethyl acetate (EA) Dichloromethane (DCM) Toluene p-Xylene m-Xylene o-Xylene
45, 59, 74 61, 70, 88 49, 84, 86 65, 91, 92 91, 105, 106 91, 105, 106 91, 105, 106
2. Experimental 2.1. Chemicals and solvents All chemicals were of reagent grade. Diethyl ether (DE), ethyl acetate (EA), dichloromethane (DCM), toluene, o-xylene, m-xylene and p-xylene were purchased from Merck (Darmstadt, Germany). Stock standard solutions were prepared in methanol at concentrations of 2000 mg L−1 for DE and EA, 250 mg L−1 for DCM and 100 mg L−1 for toluene and xylenes. Working solutions used to optimize the parameters for LPME were prepared daily at concentrations of 800 g L−1 for DE and EA, 100 g L−1 for DCM and 40 g L−1 for toluene and xylenes. Purified water was obtained from a Milli-Q water purification system (Millipore, Bedford, MA, USA). Sodium chloride (NaCl) was purchased from Showa Chemicals (Tokyo, Japan). The extraction solvents (1-octanol, benzyl alcohol, isooctane and ethylene glycol) were of analytical grade and purchased from Merck (Darmstadt, Germany). A Q 3/2 Accurel polypropylene hollow-fiber membrane (600 m i.d., 200-m wall thickness, 0.2-m pore size) was purchased from Membrana GmbH (Wuppertal, Germany). The hollow fiber was cut into 2.5-cm segments. Prior to use, the hollow fibers were cleaned by ultrasonication in acetone for 5 min. 2.2. Instrumentation All analyses were performed using an Agilent Technologies GC 6890 gas chromatography system coupled with an injection system maintained at 180 ◦ C in split mode (split ratio 10:1) and a MSD 5975 mass spectrometer. A 30-m HP-INNOWAX (0.32 mm i.d., 0.50-m film thickness) fused silica capillary column was used for separation. Helium (99.995%) was used as the carrier gas at a constant flow rate of 1.4 mL min−1 . The GC operating conditions were as follows: The initial temperature was held at 40 ◦ C for 2 min, then raised to 90 ◦ C at 25 ◦ C min−1 and held for 4.3 min, followed by increasing to 230 ◦ C at 25 ◦ C min−1 and held for 4 min. Total scan time was 17.9 min. The MS source temperature was set to 230 ◦ C. Data was acquired in selected ion monitoring (SIM) mode for quantification. The qualitative GC–MS data for the studied VOCs are shown in Table 1. 2.3. Environmental sample preparation A river sample was collected from Da-Han River (Taipei, Taiwan). The wastewater sample 1 was obtained from National Taiwan University Hospital (Taipei, Taiwan). The wastewater sample 2 was collected from Tucheng Industrial Park (New Taipei City, Taiwan). They were centrifuged at 3000 rpm for 15 min prior to analysis.
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was drawn into the microsyringe and used to pierce a headspace sample vial septum cap. The tip of the syringe inserted into one end of the hollow fiber while the other end of the fiber was fixed to the other Hamilton microsyringe (10 L, 26 gauge). A stir bar (1.5 cm ×;0.4 cm) and NaCl (30 g in 100 mL) were placed in the headspace sample vial. The septum cap was tightened and the vial was placed on the stirring hot plate. Sample solution (4 mL) was introduced into the vial using a 5-mL syringe and stirred at 1000 rpm at 95 ◦ C to achieve pre-equilibration (1 min). The extraction solvent was then pushed into the hollow fiber to impregnate the pores. After 20 min of extraction time, approximately 10 L was able to be withdrawn into a microsyringe. It usually contained 3–4 L of water at the end at the lower part. Therefore, the first 5 L was discharged from the microsyringe. The residual 5 L was injected into the GC–MS for analysis. The used fiber was discarded and a fresh one was used for the next experiment. 3. Results and discussion 3.1. Selection of extraction solvent A suitable extraction solvent is important to optimize the extraction efficiency. The polarity should match that of the target compounds, while the boiling point and volatility of the solvent should be carefully considered to prevent too much evaporation during extraction. Proper immobilization of extraction solvent in the pores of the hollow fiber is important for obtaining high extraction efficiency. Herein the properties of extraction solvents should be compatible with those of the hollow fiber. In this study, 1octanol, isooctane, ethylene glycol and benzyl alcohol were tested as the extraction solvent, at a temperature of 50 ◦ C and stirring at 500 rpm for 5 min without salt addition. As the results in Fig. 2, 1octanol shows the greatest extraction efficiency for the majority of VOCs. This may be because 1-octanol has lower polarity and viscosity than ethylene glycol and benzyl alcohol. Though isooctane has the lowest polarity and viscosity, it evaporates quickly during extraction due to its high volatility. For xylenes, m-xylene obtained higher extraction efficiency than the other two isomers. This may result from the relatively higher octanol/water partition coefficient of m-xylene (3.20) than that of p-xylene (3.15) and o-xylene (3.12). Therefore, 1-octanol was chosen as the extraction solvent for the further study. 3.2. Effect of temperature Temperature shows a significant influence on mass transfer during extraction as the temperature determines the diffusion coefficient or partition coefficient for analytes in three phases (sample matrix/headspace/hollow-fiber-supported organic phase) [29]. Considering sample matrix is water. The highest temperature was set below its boiling point (100 ◦ C). The effects of extraction temperature were studied from ambient temperature (25 ◦ C) to 95 ◦ C (Fig. 3). It was found that all analytes had the highest extraction efficiency when the temperature was 95 ◦ C. The data showed that better extraction efficiency was obtained for all analytes when using higher extraction temperature. This result may be due to the increase of Henry’s constant and diffusion coefficients of analytes in the headspace at high temperature. However, the distribution constants of analytes in the organic phase are decreased with increasing temperature [30]. In the remaining measurements, the extraction temperature was held at 95 ◦ C.
2.4. Procedures of HS-K-HFME 3.3. Effect of stirring rate A 2.5 cm hollow fiber was knotted (Fig. 1). A Hamilton syringe (50 L, 22 gauge) was used to introduce the extraction solvent to the fiber and also for GC–MS injection. A 25-L aliquot of 1-octanol
Headspace LPME is an equilibrium process in which analytes partition between the aqueous sample, the headspace and the
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P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47
Fig. 2. Extraction efficiency of different extracting solvents. The spiked concentration was 800 g L−1 for DE and EA, 100 g L−1 for DCM and 40 g L−1 for toluene and xylenes.
Fig. 3. Effect of extraction temperature. The spiked concentration was 800 g L−1 for DE and EA, 100 g L−1 for DCM and 40 g L−1 for toluene and xylenes. The extraction condition was stirring rate of 500 rpm for 5 min without salt addition.
organic extraction phase. Increasing the stirring rate enhances mass transfer in the aqueous phase and induces convection in the headspace, leading to rapid equilibrium being reached between the aqueous phase and vapor phase. The extraction was carried out at stirring rates between 0 and 1250 rpm. When the stirring rate was more than 1000 rpm, the stirring bar in the sample solution could not move steadily, disturbances were formed in the aqueous sample resulting in poorer extraction efficiency. Therefore, 1000 rpm was chosen as the appropriate stirring rate during extraction.
reduce the extraction efficiency depending on the properties of the analyte [22,31,32]. The impact of ionic strength was investigated by adding 0–30% (g/100 mL H2 O) of NaCl to the aqueous solution. The extraction efficiency reaches a maximum at 5% NaCl for toluene and xylenes but decreases with further increasing salt concentration. A possible explanation for this observation may be that the salt in the aqueous solution changed the physical properties of the Nernst diffusion layer and reduced the mass transfer rate of the analytes [33]. Nevertheless, the opposite effect was observed in ethyl acetate. Taking all analytes into consideration, the extraction was operated in the presence of 30% of NaCl.
3.4. Effect of salt 3.5. Effect of extraction time Addition of salt to the sample may have several impacts on the extraction. For SPME and LPME, a salting-out effect decreases the solubility of the analytes in the aqueous solution and enhances the partition of analytes into the adsorbent or extraction organic phase [26]. However, it has been reported that the addition of salt may also
The extraction time tested varied between 5 and 30 min (Fig. 4), with the extraction efficiency enhanced with prolonged extraction times. The amount of VOCs extracted reached a maximum after approximately 20 min. Longer times (>20 min) did not improve the
P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47
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Fig. 4. Effect of extraction time. The spiked concentration was 800 g L−1 for DE and EA, 100 g L−1 for DCM and 40 g L−1 for toluene and xylenes. The extraction condition was stirring rate of 1000 rpm at 95 ◦ C, and the addition of 30% NaCl.
Table 2 Performance of the validation analysis. Analytes
Linearity (g L−1 )
R2
LODa (g L−1 )
Enrichment factor (EF)b
RSD(%), intra-day n = 5 b
DE EA DCM Toluene p-Xylene m-Xylene o-Xylene a b c
100–80000 100–80000 10–80000 1–8000 1–8000 1–8000 1–8000
0.9967 0.9987 0.9968 0.9958 0.9957 0.9996 0.9999
10 10 5 0.3 0.2 0.3 0.3
22 29 30 70 86 85 88
RSD(%), inter-day n = 15
c
b
c
River b
Wastewater 1
Low
High
Low
High
RR (%)
RRb (%)
4.5 8.2 6.2 7.4 11.6 7.4 8.7
10.2 10.1 13.3 11.3 10.7 10.8 11.4
8.3 4.3 7.9 7.3 12.6 12.9 11.2
2.3 2.5 5.4 2.6 2.2 2.9 3.0
99.8 96.2 95.9 97.0 97.5 96.8 103.6
106.0 90.3 102.2 90.4 106.0 89.6 103.6
Limits of detection (LODs) are calculated as three times the standard deviation of three replicated runs of blank water. Amount of analytes added: 500 g L−1 for DE, EA and DCM; 50 g L−1 for toluene and xylenes. Amount of analytes added: 10,000 g L−1 for DE, EA and DCM; 1000 g L−1 for toluene and xylenes.
extraction efficiency due to significant extraction solvent evaporation and excess water vapor condensing on the fiber. Therefore, the extraction time of 20 min was chosen as the optimum. 3.6. Quantitative aspects Under optimized conditions, the linearity, LOD and precision of the method were evaluated and these results are shown in Table 2. The linearity of the method was evaluated using aqueous samples spiked with analytes at different concentrations ranging from 1 to 80,000 g L−1 . The calibration curves exhibited coefficients of determination (R2 ) ranging from 0.9957 to 0.9999 and the LOD ranged from 0.2 to 10 g L−1 . Relative standard deviations (RSDs) ranged from 4.5 to 11.6% intraday (n = 5) while inter-day (n = 15) the values were between 2.2% and 12.9%.
Table 3 Analysis of environmental water samples.
DE EA DCM Toluene p-Xylene m-Xylene o-Xylene
River water
Wastewater 1
Wastewater 2
ND ND ND ND ND ND ND
ND ND ND ND ND ND ND
ND ND 8.5 9.6 2.2 3 2.9
ND, Not detected. The value was shown as concentration (ng mL−1 ).
VOCs were 90.3–106.0% for the river sample and 95.9–103.6% for the wastewater sample 1. The results demonstrated that the method was not significantly affected by the sample matrices.
3.7. Environmental sample analysis To investigate the applicability of the method to field samples, HS-K-HFME was used to determine the levels of VOCs in three different water samples, namely a river and two wastewater specimens (Table 3). The wastewater sample 2, collected in an industrial area, was found to contain VOCs. Fig. 5 shows its chromatogram compared with those obtained from the blank wastewater samples. The river and the wastewater sample 1 were free from the target analytes at the limits of detection. They were spiked with the analytes to test matrix effect. The relative recoveries (RRs) of spiked
3.8. Method comparison Compared to other extraction techniques [13,30,34–41] shown in Table 4, without the assistance of a cooling system, the heating temperature in the study operates up to 95 ◦ C, which is the highest compared with the other methods. The required extraction time for the method was shorter than that for SPME. To protect the natural environment and the health of technicians, the use of smaller amounts (25 L) of less toxic 1-octanol is desirable.
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P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47
Fig. 5. Chromatograms of VOCs obtained by HS-K-HFME under optimized conditions (A) blank wastewater sample 1; (B) spiked wastewater sample 1 (500 g L−1 for DE, EA and DCM; 50 g L−1 for toluene and xylenes); (C) wastewater sample 2. Peaks: (a) DE; (b) EA; (c) DCM; (d) toluene; (e) p-xylene; (f) m-xylene; (g) o-xylene.
Table 4 Comparison with other extraction techniques. Method
Instrument
Analytes
Extraction solvent
Extraction time (min)
Heating temperature (◦ C)
Linearity (g L−1 )
LOD (g L−1 )
Ref.
HS-K-HFME Static HS-NTDa M-SPMEb HS-IL-SPMEc HS-SDMEd HS-SDME HS-SDME Dynamic HS-LPMEe DHS-OSFMEf HS-SPDEg MESIh
GC–MS GC–MS GC-FID GC-FID GC-FID GC-FID GC-FID GC–MS GC-FID GC–MS GC-TCD
7 VOCs 18 VOCs 12 VOCs BTEXi Trihalomethanes 2-Butoxylethanol Methyl t-butyl ether 9 Alcohols BTEX 3 ethers, 9 alcohols BTEX, chloroform
1-Octanol – – [C6 MIM][PF6 ] Benzyl alcohol Benzyl alcohol Benzyl alcohol 1-Octanol 1-Dodecane – –
20 30–60 30 30 10 15 7.5 9.5 5.4 –
Contents lists available at ScienceDirect
Journal of Chromatography A journal homepage: www.elsevier.com/locate/chroma
Determination of volatile organic compounds in water using headspace knotted hollow fiber microextraction Pai-Shan Chen a,b,∗ , Yu-Hsiang Tseng a , Yuh-Lin Chuang a , Jung-Hsuan Chen a,b a b
Department and Graduate Institute of Forensic Medicine, National Taiwan University, Taipei 10002, Taiwan Forensic and Clinical Toxicology Center, National Taiwan University College of Medicine and National Taiwan University Hospital, Taiwan
a r t i c l e
i n f o
Article history: Received 10 January 2015 Received in revised form 24 March 2015 Accepted 24 March 2015 Available online 31 March 2015 Keywords: Headspace Hollow fiber microextraction Volatile organic compounds Gas chromatography–mass spectrometry River water Wastewater
a b s t r a c t An efficient and effective headspace microextraction technique named static headspace knotted hollow fiber microextraction (HS-K-HFME) has been developed for the determination of volatile organic compounds (VOCs) in water samples. The knot-shaped hollow fiber is filled with 25 L of the extraction solvent. The excess solvent forms a large droplet (13 L) and is held in the center of the knot. Even after 20 min of extraction time at high temperature (95 ◦ C) without cooling, there was still enough volume of extraction solvent for gas chromatography–mass spectrometry (GC–MS) analysis, which extends the choice of solvents for headspace LPME. Moreover, the knot-shaped fiber has a larger extraction contact interface, which increases the rate of mass transfer between the headspace and extraction solvent film attached to the fiber, thus improving the extraction efficiency. The effects of extraction solvent, temperature, stirring rate, salt concentration and extraction time on extraction performance were optimized. The calibration curves exhibited coefficients of determination (R2 ) ranging from 0.9957 to 0.9999 and the limit of detection (LOD) ranged from 0.2 to 10 g L−1 . Relative standard deviations (RSDs) ranged from 4.5% to 11.6% for intraday measurements (n = 5). Interday (n = 15) values were between 2.2% and 12.9%. The relative recoveries (RRs) ranged from 90.3% to 106.0% for river water and 95.9% to 103.6% for wastewater. © 2015 Elsevier B.V. All rights reserved.
1. Introduction Volatile organic compounds (VOCs) are defined as having a boiling point that ranges between 50 ◦ C and 260 ◦ C [1]. VOCs are widely emitted from sources such as paints, varnishes, solvents and preservatives, and become major environmental pollutants because of their high volatility and toxicity [2,3]. Due to their physical properties, VOCs can easily cross lipid membranes and distribute to well-perfused organs; exposure to VOCs may thus result in both acute and chronic health effects. Diethyl ether (DE) and ethyl acetate (EA) are two common VOCs, which are used worldwide in flavoring and perfumery and in smokeless power manufacture. They are mildly irritating to eyes, nose and throat. Exposure to high levels may cause dizziness or to pass out. Especially for DE, severe over exposure may result in death [4]. Certain VOCs are more toxic and persistent in water, soil, and organisms, such as dichloromethane (DCM), toluene, o-xylene, m-xylene, and
∗ Corresponding author at: Department and Graduate Institute of Forensic Medicine, National Taiwan University, Taipei 10002, Taiwan. Tel.: +886 2 2312 3456x65495; fax: +886 2 2321 8438. E-mail address: [email protected] (P.-S. Chen). http://dx.doi.org/10.1016/j.chroma.2015.03.067 0021-9673/© 2015 Elsevier B.V. All rights reserved.
p-xylene. They are considered one of the major causes of environmental pollution because of widespread occurrences of leakage from underground gasoline storage tanks and spills. Long term exposure of them has been associated with liver and kidney damage, intestinal tract disturbances and central nervous system depression [4–6]. Therefore, the United States Environmental Protection Agency (EPA) has regulated 0.005 and 10 mg L−1 as the maximum permissible contaminant level for dichloromethane and total xylene respectively in drinking water [7]. As a result of this low limit, it is necessary to develop highly sensitive and efficient analytical methods to detect VOCs in the aquatic environment. Analyte extraction and pretreatment is the most challenging and time-consuming step in most chromatographic procedures. In the last few years, research trends in separation science have oriented toward minimizing the sample pretreatment steps. Solidphase microextraction (SPME) and liquid-phase microextraction (LPME) have been developed based on this concept. SPME was first introduced by Pawliszyn and coworkers [8] and involves the partition of analytes between sample matrices and a polymercoated stationary phase on a silica fiber. It enables the extraction and simultaneous preconcentration of analytes from aqueous samples. There are three major types of SPME: direct-immersion [9], headspace (HS) [10–12] and membrane-protected SPME [13].
42
P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47
Fig. 1. The extraction equipment of HS-K-HFME.
Among them, HS-SPME analysis is the most common technique for the determination of VOCs as it significantly reduces the interference from dirty matrices, which is generally associated with direct-immersion SPME [14–16]. Lately, LPME has emerged as an attractive alternative to conventional liquid–liquid extraction (LLE). Jeannot and Cantwell proposed single-drop microextraction (SDME) in which an 8-L droplet of organic solvent was suspended at the end of a Teflon rod and immersed in an aqueous solution to extract analyte specimens [17]. An improvement involved the use of a microsyringe to hold the solvent drop [18]. To increase the rate of mass transfer, Lee et al. developed a dynamic liquid phase microextraction technique using a short hollow fiber inserted onto a microsyringe [19,20]. The hydrophobic and porous fiber was filled with organic solvent, which stabilizes the solvent successfully in comparison with holding a droplet at the tip of the microsyringe [21–23]. The immersion mode yields high enrichment factors, high sensitivity and ruggedness. However, headspace LPME using a hollow fiber presents more challenges than other types of LPME because the extraction solvents compatible with gas chromatography (GC) usually evaporate quickly in the headspace. For headspace LPME, SDME and fiber-protected LPME have been developed. Theis et al. [24] introduced headspace SDME using 1octanol as the extraction solvent for analyzing VOCs, which has a relatively low vapor pressure. Shen and Lee [25] used dynamic headspace LPME to improve the stability of headspace SDME. A thin organic solvent film was formed on the inner syringe wall. The dynamic operation increased and refreshed the interface between the gas phase and the extraction solvent, thus increasing the extraction efficiency significantly. Jiang et al. [26] developed a dynamic hollow fiber-supported headspace liquid-phase microextraction (DHF-HS-LPME) technique. A hollow fiber filled with the solvent was inserted onto a microsyringe needle which was kept vertically in the headspace of the sample solution. The solvent within the fiber could be moved using a programmable syringe pump. This method enabled the use of higher solvent volumes, increased the extraction contact interface and stabilized the organic solvent within the fiber. In such analyses, headspace gas is usually injected into a GC directly. A major concern is that the organic solvents commonly used in GC have high vapor pressures, which result in quick evaporation in the headspace and limits the possible selection
of extraction solvents. In 2007, Huang and Huang [21] applied a solvent cooling system to the microsyringe which decreased the temperature of the solvent when it was drawn into the syringe. This decreased temperature minimized the loss of extraction solvent but also caused slow equilibration and led to longer extraction times. Chen et al. [27] developed a dynamic hook-type liquid-phase microextraction (DHT-LPME). The extraction solvent was easily and completely withdrawn into the microsyringe, while the hook shape removed interference from air bubbles which commonly persist when the hollow fiber is placed vertically. Subsequently the same group introduced dynamic headspace time-extended helix LPME (DHS-TEH-LPME) to overcome the slow equilibrium caused by cooling system. An aluminum heating block was used to heat the sample vial while the extraction solvent was condensed using a cooling system. The extraction temperature up to 80 ◦ C was used for 60 min. Chen et al. [28] developed a PTFE vial cap with a cylindrical cavity to hold a 40-L droplet of volatile extraction solvents, such as acetone, in the headspace. The cooling system, based on a thermoelectric cooler (TEC), was used to lower the temperature of the vial cap below zero. This method enabled a limit of detection (LOD) for chlorobenzenes between 4 and 8 g L−1 . It is worth noting that, in order to extend the selection of solvents and reduce solvent loss during headspace, LPME extraction processes, a cooling system is usually required. However, this also results in slow equilibrium and longer extraction times to obtain satisfactory extraction efficiency. To address these issues, we proposed a simple and efficient headspace LPME technique named headspace knotted hollow fiber microextraction (HS-K-HFME) for the determination of VOCs in water samples. A 2.5 cm long hollow fiber was bent into a figure eight knot shape (Fig. 1) and filled with 25 L of extraction solvent. AA 2.5 cm fiber can hold 12 L of solvent approximately. The excess solvent which was not held inside the fiber formed a large droplet (approximately 13 L). The droplet was held steadily at the center of the knot. As a result of the figure eight shape, there is no interference from air bubbles in the fiber. The extraction solvent is withdrawn into the microsyringe easily and the long fiber has a large contact surface that increases the extraction interface between the headspace and the organic solvent. This large volume is sufficient to compensate for evaporative losses during sampling. Moreover, during the extraction of VOCs, the specimen solution can also be heated.
P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47 Table 1 Qualitative GC–MS data for the studied VOCs. Analyte
Quantitative and confirmed ions (m/z)
Diethyl ether (DE) Ethyl acetate (EA) Dichloromethane (DCM) Toluene p-Xylene m-Xylene o-Xylene
45, 59, 74 61, 70, 88 49, 84, 86 65, 91, 92 91, 105, 106 91, 105, 106 91, 105, 106
2. Experimental 2.1. Chemicals and solvents All chemicals were of reagent grade. Diethyl ether (DE), ethyl acetate (EA), dichloromethane (DCM), toluene, o-xylene, m-xylene and p-xylene were purchased from Merck (Darmstadt, Germany). Stock standard solutions were prepared in methanol at concentrations of 2000 mg L−1 for DE and EA, 250 mg L−1 for DCM and 100 mg L−1 for toluene and xylenes. Working solutions used to optimize the parameters for LPME were prepared daily at concentrations of 800 g L−1 for DE and EA, 100 g L−1 for DCM and 40 g L−1 for toluene and xylenes. Purified water was obtained from a Milli-Q water purification system (Millipore, Bedford, MA, USA). Sodium chloride (NaCl) was purchased from Showa Chemicals (Tokyo, Japan). The extraction solvents (1-octanol, benzyl alcohol, isooctane and ethylene glycol) were of analytical grade and purchased from Merck (Darmstadt, Germany). A Q 3/2 Accurel polypropylene hollow-fiber membrane (600 m i.d., 200-m wall thickness, 0.2-m pore size) was purchased from Membrana GmbH (Wuppertal, Germany). The hollow fiber was cut into 2.5-cm segments. Prior to use, the hollow fibers were cleaned by ultrasonication in acetone for 5 min. 2.2. Instrumentation All analyses were performed using an Agilent Technologies GC 6890 gas chromatography system coupled with an injection system maintained at 180 ◦ C in split mode (split ratio 10:1) and a MSD 5975 mass spectrometer. A 30-m HP-INNOWAX (0.32 mm i.d., 0.50-m film thickness) fused silica capillary column was used for separation. Helium (99.995%) was used as the carrier gas at a constant flow rate of 1.4 mL min−1 . The GC operating conditions were as follows: The initial temperature was held at 40 ◦ C for 2 min, then raised to 90 ◦ C at 25 ◦ C min−1 and held for 4.3 min, followed by increasing to 230 ◦ C at 25 ◦ C min−1 and held for 4 min. Total scan time was 17.9 min. The MS source temperature was set to 230 ◦ C. Data was acquired in selected ion monitoring (SIM) mode for quantification. The qualitative GC–MS data for the studied VOCs are shown in Table 1. 2.3. Environmental sample preparation A river sample was collected from Da-Han River (Taipei, Taiwan). The wastewater sample 1 was obtained from National Taiwan University Hospital (Taipei, Taiwan). The wastewater sample 2 was collected from Tucheng Industrial Park (New Taipei City, Taiwan). They were centrifuged at 3000 rpm for 15 min prior to analysis.
43
was drawn into the microsyringe and used to pierce a headspace sample vial septum cap. The tip of the syringe inserted into one end of the hollow fiber while the other end of the fiber was fixed to the other Hamilton microsyringe (10 L, 26 gauge). A stir bar (1.5 cm ×;0.4 cm) and NaCl (30 g in 100 mL) were placed in the headspace sample vial. The septum cap was tightened and the vial was placed on the stirring hot plate. Sample solution (4 mL) was introduced into the vial using a 5-mL syringe and stirred at 1000 rpm at 95 ◦ C to achieve pre-equilibration (1 min). The extraction solvent was then pushed into the hollow fiber to impregnate the pores. After 20 min of extraction time, approximately 10 L was able to be withdrawn into a microsyringe. It usually contained 3–4 L of water at the end at the lower part. Therefore, the first 5 L was discharged from the microsyringe. The residual 5 L was injected into the GC–MS for analysis. The used fiber was discarded and a fresh one was used for the next experiment. 3. Results and discussion 3.1. Selection of extraction solvent A suitable extraction solvent is important to optimize the extraction efficiency. The polarity should match that of the target compounds, while the boiling point and volatility of the solvent should be carefully considered to prevent too much evaporation during extraction. Proper immobilization of extraction solvent in the pores of the hollow fiber is important for obtaining high extraction efficiency. Herein the properties of extraction solvents should be compatible with those of the hollow fiber. In this study, 1octanol, isooctane, ethylene glycol and benzyl alcohol were tested as the extraction solvent, at a temperature of 50 ◦ C and stirring at 500 rpm for 5 min without salt addition. As the results in Fig. 2, 1octanol shows the greatest extraction efficiency for the majority of VOCs. This may be because 1-octanol has lower polarity and viscosity than ethylene glycol and benzyl alcohol. Though isooctane has the lowest polarity and viscosity, it evaporates quickly during extraction due to its high volatility. For xylenes, m-xylene obtained higher extraction efficiency than the other two isomers. This may result from the relatively higher octanol/water partition coefficient of m-xylene (3.20) than that of p-xylene (3.15) and o-xylene (3.12). Therefore, 1-octanol was chosen as the extraction solvent for the further study. 3.2. Effect of temperature Temperature shows a significant influence on mass transfer during extraction as the temperature determines the diffusion coefficient or partition coefficient for analytes in three phases (sample matrix/headspace/hollow-fiber-supported organic phase) [29]. Considering sample matrix is water. The highest temperature was set below its boiling point (100 ◦ C). The effects of extraction temperature were studied from ambient temperature (25 ◦ C) to 95 ◦ C (Fig. 3). It was found that all analytes had the highest extraction efficiency when the temperature was 95 ◦ C. The data showed that better extraction efficiency was obtained for all analytes when using higher extraction temperature. This result may be due to the increase of Henry’s constant and diffusion coefficients of analytes in the headspace at high temperature. However, the distribution constants of analytes in the organic phase are decreased with increasing temperature [30]. In the remaining measurements, the extraction temperature was held at 95 ◦ C.
2.4. Procedures of HS-K-HFME 3.3. Effect of stirring rate A 2.5 cm hollow fiber was knotted (Fig. 1). A Hamilton syringe (50 L, 22 gauge) was used to introduce the extraction solvent to the fiber and also for GC–MS injection. A 25-L aliquot of 1-octanol
Headspace LPME is an equilibrium process in which analytes partition between the aqueous sample, the headspace and the
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P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47
Fig. 2. Extraction efficiency of different extracting solvents. The spiked concentration was 800 g L−1 for DE and EA, 100 g L−1 for DCM and 40 g L−1 for toluene and xylenes.
Fig. 3. Effect of extraction temperature. The spiked concentration was 800 g L−1 for DE and EA, 100 g L−1 for DCM and 40 g L−1 for toluene and xylenes. The extraction condition was stirring rate of 500 rpm for 5 min without salt addition.
organic extraction phase. Increasing the stirring rate enhances mass transfer in the aqueous phase and induces convection in the headspace, leading to rapid equilibrium being reached between the aqueous phase and vapor phase. The extraction was carried out at stirring rates between 0 and 1250 rpm. When the stirring rate was more than 1000 rpm, the stirring bar in the sample solution could not move steadily, disturbances were formed in the aqueous sample resulting in poorer extraction efficiency. Therefore, 1000 rpm was chosen as the appropriate stirring rate during extraction.
reduce the extraction efficiency depending on the properties of the analyte [22,31,32]. The impact of ionic strength was investigated by adding 0–30% (g/100 mL H2 O) of NaCl to the aqueous solution. The extraction efficiency reaches a maximum at 5% NaCl for toluene and xylenes but decreases with further increasing salt concentration. A possible explanation for this observation may be that the salt in the aqueous solution changed the physical properties of the Nernst diffusion layer and reduced the mass transfer rate of the analytes [33]. Nevertheless, the opposite effect was observed in ethyl acetate. Taking all analytes into consideration, the extraction was operated in the presence of 30% of NaCl.
3.4. Effect of salt 3.5. Effect of extraction time Addition of salt to the sample may have several impacts on the extraction. For SPME and LPME, a salting-out effect decreases the solubility of the analytes in the aqueous solution and enhances the partition of analytes into the adsorbent or extraction organic phase [26]. However, it has been reported that the addition of salt may also
The extraction time tested varied between 5 and 30 min (Fig. 4), with the extraction efficiency enhanced with prolonged extraction times. The amount of VOCs extracted reached a maximum after approximately 20 min. Longer times (>20 min) did not improve the
P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47
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Fig. 4. Effect of extraction time. The spiked concentration was 800 g L−1 for DE and EA, 100 g L−1 for DCM and 40 g L−1 for toluene and xylenes. The extraction condition was stirring rate of 1000 rpm at 95 ◦ C, and the addition of 30% NaCl.
Table 2 Performance of the validation analysis. Analytes
Linearity (g L−1 )
R2
LODa (g L−1 )
Enrichment factor (EF)b
RSD(%), intra-day n = 5 b
DE EA DCM Toluene p-Xylene m-Xylene o-Xylene a b c
100–80000 100–80000 10–80000 1–8000 1–8000 1–8000 1–8000
0.9967 0.9987 0.9968 0.9958 0.9957 0.9996 0.9999
10 10 5 0.3 0.2 0.3 0.3
22 29 30 70 86 85 88
RSD(%), inter-day n = 15
c
b
c
River b
Wastewater 1
Low
High
Low
High
RR (%)
RRb (%)
4.5 8.2 6.2 7.4 11.6 7.4 8.7
10.2 10.1 13.3 11.3 10.7 10.8 11.4
8.3 4.3 7.9 7.3 12.6 12.9 11.2
2.3 2.5 5.4 2.6 2.2 2.9 3.0
99.8 96.2 95.9 97.0 97.5 96.8 103.6
106.0 90.3 102.2 90.4 106.0 89.6 103.6
Limits of detection (LODs) are calculated as three times the standard deviation of three replicated runs of blank water. Amount of analytes added: 500 g L−1 for DE, EA and DCM; 50 g L−1 for toluene and xylenes. Amount of analytes added: 10,000 g L−1 for DE, EA and DCM; 1000 g L−1 for toluene and xylenes.
extraction efficiency due to significant extraction solvent evaporation and excess water vapor condensing on the fiber. Therefore, the extraction time of 20 min was chosen as the optimum. 3.6. Quantitative aspects Under optimized conditions, the linearity, LOD and precision of the method were evaluated and these results are shown in Table 2. The linearity of the method was evaluated using aqueous samples spiked with analytes at different concentrations ranging from 1 to 80,000 g L−1 . The calibration curves exhibited coefficients of determination (R2 ) ranging from 0.9957 to 0.9999 and the LOD ranged from 0.2 to 10 g L−1 . Relative standard deviations (RSDs) ranged from 4.5 to 11.6% intraday (n = 5) while inter-day (n = 15) the values were between 2.2% and 12.9%.
Table 3 Analysis of environmental water samples.
DE EA DCM Toluene p-Xylene m-Xylene o-Xylene
River water
Wastewater 1
Wastewater 2
ND ND ND ND ND ND ND
ND ND ND ND ND ND ND
ND ND 8.5 9.6 2.2 3 2.9
ND, Not detected. The value was shown as concentration (ng mL−1 ).
VOCs were 90.3–106.0% for the river sample and 95.9–103.6% for the wastewater sample 1. The results demonstrated that the method was not significantly affected by the sample matrices.
3.7. Environmental sample analysis To investigate the applicability of the method to field samples, HS-K-HFME was used to determine the levels of VOCs in three different water samples, namely a river and two wastewater specimens (Table 3). The wastewater sample 2, collected in an industrial area, was found to contain VOCs. Fig. 5 shows its chromatogram compared with those obtained from the blank wastewater samples. The river and the wastewater sample 1 were free from the target analytes at the limits of detection. They were spiked with the analytes to test matrix effect. The relative recoveries (RRs) of spiked
3.8. Method comparison Compared to other extraction techniques [13,30,34–41] shown in Table 4, without the assistance of a cooling system, the heating temperature in the study operates up to 95 ◦ C, which is the highest compared with the other methods. The required extraction time for the method was shorter than that for SPME. To protect the natural environment and the health of technicians, the use of smaller amounts (25 L) of less toxic 1-octanol is desirable.
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P.-S. Chen et al. / J. Chromatogr. A 1395 (2015) 41–47
Fig. 5. Chromatograms of VOCs obtained by HS-K-HFME under optimized conditions (A) blank wastewater sample 1; (B) spiked wastewater sample 1 (500 g L−1 for DE, EA and DCM; 50 g L−1 for toluene and xylenes); (C) wastewater sample 2. Peaks: (a) DE; (b) EA; (c) DCM; (d) toluene; (e) p-xylene; (f) m-xylene; (g) o-xylene.
Table 4 Comparison with other extraction techniques. Method
Instrument
Analytes
Extraction solvent
Extraction time (min)
Heating temperature (◦ C)
Linearity (g L−1 )
LOD (g L−1 )
Ref.
HS-K-HFME Static HS-NTDa M-SPMEb HS-IL-SPMEc HS-SDMEd HS-SDME HS-SDME Dynamic HS-LPMEe DHS-OSFMEf HS-SPDEg MESIh
GC–MS GC–MS GC-FID GC-FID GC-FID GC-FID GC-FID GC–MS GC-FID GC–MS GC-TCD
7 VOCs 18 VOCs 12 VOCs BTEXi Trihalomethanes 2-Butoxylethanol Methyl t-butyl ether 9 Alcohols BTEX 3 ethers, 9 alcohols BTEX, chloroform
1-Octanol – – [C6 MIM][PF6 ] Benzyl alcohol Benzyl alcohol Benzyl alcohol 1-Octanol 1-Dodecane – –
20 30–60 30 30 10 15 7.5 9.5 5.4 –
